Based on promising preclinical results, several different TGF-beta pathway antagonists are in early phase clinical trials for the treatment of advanced cancer. However, given the complex biology of TGF-beta, the successful development of TGF-beta antagonists for cancer therapy will depend on a clear understanding of how these agents work, and the related question of how to select patients who will benefit from this type of treatment. Using a panel of 12 mouse syngeneic allograft models of metastatic breast cancer, with metastatic burden as the primary endpoint, we uncovered heterogeneous responses to TGF-beta antagonism. TGF-beta pathway blockade inhibited metastasis in some models, while having no effect on or stimulating metastasis in other models. We are continuing to apply targeted and discovery-based approaches to address molecular and biological mechanisms underlying the heterogeneity of therapeutic response and to generate useful predictive biomarkers. Unexpectedly, neither TGF-beta expression levels nor extent of TGF-beta pathway activation in the primary tumor predict response to anti-TGF-beta therapy. However, tumors from models showing a desirable response to TGF-beta antagonism are characterized by transcriptomic evidence of TGF-beta pathway activation in the untreated state. Thus in vivo, the output of TGF-beta signaling may be a more sensitive indicator of the activation status of the TGF-beta pathway than any of the input parameters such as ligand levels and status of signal transduction components. We are generating gene expression signatures that might predict response to anti-TGF-beta therapy and will test these in additional preclinical models. We have also identified candidate molecular determinants of the therapeutic response to TGF-beta and are testing these by knockdown and overexpression approaches in the tumor models. We are also continuing to address ways of improving the therapeutic efficacy of TGF-beta antagonism. Correlative clinical data and literature evidence suggest that whereas TGF-beta1 is primarily associated with poor outcome in many cancers, TGF-beta3 may actually oppose TGF-beta1 and be associated with good outcome. We have optimized a method for accurate quantitation of TGF-beta protein isoform expression and have shown that the mammary gland is one of the few adult organs to express significant levels of TGF-beta3. We find TGF-beta1 to be consistently upregulated and TGF-beta3 to be downregulated when compared with the normal mammary gland. Our preliminary data with TGF-beta isoform-selective antibodies suggests that there is no advantage to sparing TGF-beta3 in terms of therapeutic efficacy, but that there may be positive metabolic consequences of this approach. Our integrated genomic approaches are also suggesting drugs that could be combined with TGF-beta antagonists to improve therapeutic efficacy, and we are actively pursuing the top leads in our preclinical models.